Twelve of fourteen strains of cultured skin fibroblasts from patients with I-cell disease (mucolipidosis II) have been found to possess abnormally high intracellular levels of the amino acid cystine and in this respect bear a striking resemblance to the characteristic abnormality encountered in human cystinosis. In contrast to these results nine of eleven human cell lines from patiens with the biochemically similar, but clinically less severe, disorder known as mucolipidosis III were essentially normal with respect to levels of this amino acid while two strains exhibited moderately enhanced levels of cystine. Comparative studies of the intracellular localization of excess cystine in cystinotic and I-cell fibroblasts showed that in both cases the amino acid was associated predominately with the lysosomal fraction of these cell types. These observations support the notion that human cystinosis is due to some (as yet unknown) lysosomal dysfunction.